鸽A群轮状病毒实时荧光RT-PCR检测方法的建立及应用  

Establishment and Application of a Fluorescent RT-PCR for Detecting Pigeon Group A Rotavirus

作　　者：邢安琪 彭真奇 蒋文明[1] 克军宏 李阳 陈璐 王静静[1] 许立华[2] 于晓慧 刘华雷[1] Xing Anqi;Peng Zhenqi;Jiang Wenming;Ke Junhong;Li Yang;Chen Lu;Wang Jingjing;Xu Lihua;Yu Xiaohui;Liu Hualei(China Animal Health and Epidemiology Center,Qingdao,Shandong 266032,China;Ningxia University,Yinchuan,Ningxia 750021,China;Anhui Agricultural University,Hefei,Anhui 230036,China;Tarim University,Alaer,Xinjiang 843300,China;Shandong Agricultural University,Tai'an,Shandong 271018,China)

机构地区：[1]中国动物卫生与流行病学中心,山东青岛266032 [2]宁夏大学,宁夏银川750021 [3]安徽农业大学,安徽合肥230036 [4]塔里木大学,新疆阿拉尔843300 [5]山东农业大学,山东泰安271018

出　　处：《中国动物检疫》2023年第9期91-97,共7页China Animal Health Inspection

基　　金：中央引导地方科技发展资金项目;国家重点研发计划项目(2016YFD0500800)。

摘　　要：鸽A群轮状病毒(group A rotavirus,RVA)是影响养鸽业健康发展的主要病原之一。为实现鸽RVA的高通量快速检测,根据目前国内外流行的鸽RVA VP6基因保守区域设计特异性引物与探针,建立了一种可以特异性检测鸽RVA的实时荧光RT-PCR方法,并对该方法的灵敏度、特异性和重复性进行了评估。结果显示,该方法灵敏度高,最低检测限为7.22×10^(2)copies/μL;特异性强,与鸽群其他常见病毒无交叉反应;重复性好,组内与组间重复变异系数均小于1.5%。利用该方法和普通RT-PCR方法对200份临床样品进行检测,发现荧光RT-PCR方法的病毒阳性检出率高于普通RT-PCR方法,二者符合率为94.00%。结果表明,本研究建立的实时荧光RT-PCR方法敏感、特异、准确,重复性好,可为开展鸽RVA的临床检测及流行病学调查提供技术支持。Pigeon group A rotavirus(RVA)is one of major pathogens affecting healthy development of pigeon industry.In order to rapidly detect RVA at high throughput,specific primers and probes were designed according to the conserved region of pigeon RVA VP6 gene currently prevalent worldwide,a real-time fluorescent RT-PCR was established,followed by evaluation on its sensitivity,specificity and repeatability.The results showed that its lowest detection limit was 7.22×10^(2) copies/μl due to its high sensitivity;no cross-reactivity with other common viruses in pigeons was observed due to its strong specificity;the intra/inter group repeated coefficients of variation were both less than 2%due to its good repeatability.200 clinical samples were detected by the established method and conventional RT-PCR,the positive detection rate by the former was higher than the latter,with a coincidence rate of 94.00%.In conclusion,the established method was sensitive,specific,accurate and repeatable,which could provide technical support for clinical detection and epidemiological investigation of pigeon RVA.

关 键 词：鸽A群轮状病毒 实时荧光RT-PCR 性能评估 应用 

分 类 号：S855.3[农业科学—临床兽医学]